Bottom Line:
Surprisingly, the upregulated expression of the ubiquitin ligases atrogin-1 and MuRF1 was not influenced by curcumin.Results suggest that sepsis-induced muscle proteolysis can be blocked by curcumin and that this effect may, at least in part, be caused by inhibited NF-kappaB and p38 activities.The results also suggest that there is not an absolute correlation between changes in muscle protein breakdown rates and changes in atrogin-1 and MuRF1 expression during treatment of muscle wasting.

ABSTRACTWe tested the hypothesis that treatment of rats with curcumin prevents sepsis-induced muscle protein degradation. In addition, we determined the influence of curcumin on different proteolytic pathways that are activated in septic muscle (i.e., ubiquitin-proteasome-, calpain-, and cathepsin L-dependent proteolysis) and examined the role of NF-kappaB and p38/MAP kinase inactivation in curcumin-induced inhibition of muscle protein breakdown. Rats were made septic by cecal ligation and puncture or were sham-operated. Groups of rats were treated with three intraperitoneal doses (600 mg/kg) of curcumin or corresponding volumes of solvent. Protein breakdown rates were measured as release of tyrosine from incubated extensor digitorum longus muscles. Treatment with curcumin prevented sepsis-induced increase in muscle protein breakdown. Surprisingly, the upregulated expression of the ubiquitin ligases atrogin-1 and MuRF1 was not influenced by curcumin. When muscles from septic rats were treated with curcumin in vitro, proteasome-, calpain-, and cathepsin L-dependent protein breakdown rates were reduced, and nuclear NF-kappaB/p65 expression and activity as well as levels of phosphorylated (activated) p38 were decreased. Results suggest that sepsis-induced muscle proteolysis can be blocked by curcumin and that this effect may, at least in part, be caused by inhibited NF-kappaB and p38 activities. The results also suggest that there is not an absolute correlation between changes in muscle protein breakdown rates and changes in atrogin-1 and MuRF1 expression during treatment of muscle wasting.

fig7: (a) p65 levels determined by Western blotting and (b) p65 activity in the nuclear fraction of extensor digitorum longus muscles from septic rats incubated for 2 hours in the absence or presence of 100 μM curcumin.Results are means ± SEM with n = 4 in each group. AU: arbitrary units. *P < .05 versus control by Student’s t-test. (c) The effect of the p65 inhibitor PTD-p65-P1 on protein breakdown rates in incubated muscles from septic rats. Muscles were incubated in the presence of 100 μM PTD-p65-P1 or 100 μM of the inactive peptide PTD as control. Results are means ± SEM with n = 7 in each group. *P < .05 versus PTD by Student’s t-test.

Mentions:
Because treatment of septic rats with curcumin in vivo reduced NF-κB/p65 activity in skeletal muscle (Figure 4), we tested whether a similar mechanism may be involved in the direct effect of curcumin in septic muscle. When muscles from septic rats were incubated in the presence of 100 μM curcumin, the nuclear levels of p65 decreased(Figure 7(a)) and p65 DNA binding activity was reduced (Figure 7(b)). In additionalexperiments, treatment of incubated muscles from sham-operated rats did notsignificantly influence p65 activity (p65 activity was 95 ± 8% ofcontrol value in muscles from sham-operated rats treated with 100 μM curcumin in vitro for 2 hours; n = 4 for both control and curcumin-treated muscles). The results in Figures 7(a) and 7(b) suggest that the inhibition of protein breakdown caused by treatment of incubated muscles from septic rats with curcumin may at least in part reflect inhibited NF-κB/p65 activity. In order to further test the potential role of NF-κBinhibition on protein breakdown, muscles from septic rats were incubated in the presence of 100 μM of the specific p65 inhibitor PTD-p65-P1 [43]. This inhibitor consists of a syntheticp65 peptide containing the serine 276 phosphorylation site and linked with a protein transduction site (PTD), a short protein sequence that can enter cells to deliver its cargo without any receptor [43]. Control muscles were treated with 100 μM PTD thatwas not linked to another peptide. Treatment of the muscles with PTD-p65-P1resulted in an approximately 30% reduction of protein breakdown (Figure 7(c)).

fig7: (a) p65 levels determined by Western blotting and (b) p65 activity in the nuclear fraction of extensor digitorum longus muscles from septic rats incubated for 2 hours in the absence or presence of 100 μM curcumin.Results are means ± SEM with n = 4 in each group. AU: arbitrary units. *P < .05 versus control by Student’s t-test. (c) The effect of the p65 inhibitor PTD-p65-P1 on protein breakdown rates in incubated muscles from septic rats. Muscles were incubated in the presence of 100 μM PTD-p65-P1 or 100 μM of the inactive peptide PTD as control. Results are means ± SEM with n = 7 in each group. *P < .05 versus PTD by Student’s t-test.

Mentions:
Because treatment of septic rats with curcumin in vivo reduced NF-κB/p65 activity in skeletal muscle (Figure 4), we tested whether a similar mechanism may be involved in the direct effect of curcumin in septic muscle. When muscles from septic rats were incubated in the presence of 100 μM curcumin, the nuclear levels of p65 decreased(Figure 7(a)) and p65 DNA binding activity was reduced (Figure 7(b)). In additionalexperiments, treatment of incubated muscles from sham-operated rats did notsignificantly influence p65 activity (p65 activity was 95 ± 8% ofcontrol value in muscles from sham-operated rats treated with 100 μM curcumin in vitro for 2 hours; n = 4 for both control and curcumin-treated muscles). The results in Figures 7(a) and 7(b) suggest that the inhibition of protein breakdown caused by treatment of incubated muscles from septic rats with curcumin may at least in part reflect inhibited NF-κB/p65 activity. In order to further test the potential role of NF-κBinhibition on protein breakdown, muscles from septic rats were incubated in the presence of 100 μM of the specific p65 inhibitor PTD-p65-P1 [43]. This inhibitor consists of a syntheticp65 peptide containing the serine 276 phosphorylation site and linked with a protein transduction site (PTD), a short protein sequence that can enter cells to deliver its cargo without any receptor [43]. Control muscles were treated with 100 μM PTD thatwas not linked to another peptide. Treatment of the muscles with PTD-p65-P1resulted in an approximately 30% reduction of protein breakdown (Figure 7(c)).

Bottom Line:
Surprisingly, the upregulated expression of the ubiquitin ligases atrogin-1 and MuRF1 was not influenced by curcumin.Results suggest that sepsis-induced muscle proteolysis can be blocked by curcumin and that this effect may, at least in part, be caused by inhibited NF-kappaB and p38 activities.The results also suggest that there is not an absolute correlation between changes in muscle protein breakdown rates and changes in atrogin-1 and MuRF1 expression during treatment of muscle wasting.

ABSTRACTWe tested the hypothesis that treatment of rats with curcumin prevents sepsis-induced muscle protein degradation. In addition, we determined the influence of curcumin on different proteolytic pathways that are activated in septic muscle (i.e., ubiquitin-proteasome-, calpain-, and cathepsin L-dependent proteolysis) and examined the role of NF-kappaB and p38/MAP kinase inactivation in curcumin-induced inhibition of muscle protein breakdown. Rats were made septic by cecal ligation and puncture or were sham-operated. Groups of rats were treated with three intraperitoneal doses (600 mg/kg) of curcumin or corresponding volumes of solvent. Protein breakdown rates were measured as release of tyrosine from incubated extensor digitorum longus muscles. Treatment with curcumin prevented sepsis-induced increase in muscle protein breakdown. Surprisingly, the upregulated expression of the ubiquitin ligases atrogin-1 and MuRF1 was not influenced by curcumin. When muscles from septic rats were treated with curcumin in vitro, proteasome-, calpain-, and cathepsin L-dependent protein breakdown rates were reduced, and nuclear NF-kappaB/p65 expression and activity as well as levels of phosphorylated (activated) p38 were decreased. Results suggest that sepsis-induced muscle proteolysis can be blocked by curcumin and that this effect may, at least in part, be caused by inhibited NF-kappaB and p38 activities. The results also suggest that there is not an absolute correlation between changes in muscle protein breakdown rates and changes in atrogin-1 and MuRF1 expression during treatment of muscle wasting.